Ethanol itself promotes the epithelial-mesenchymal transition of CRC cells via the TGF-β/Smad/Snail axis. (A) An ELISA was used to detect the expression of extracellular TGF-β1 in cells treated with ethanol. (B) An ELISA was used to detect the expression of intracellular and extracellular TGF-β1 in cells treated with acetaldehyde. (C) Representative western blots for TGF-β signalling-associated proteins. The values under the membrane represent the expression of genes normalized to the expression of the reference gene GAPDH. “E-ca”, “β-cat”, “FN”, “N-ca”, and “Vim” indicate E-cadherin, β-catenin, fibronectin, N-cadherin and vimentin, respectively. (D) Immunofluorescence assay of p-Smad2 protein in ethanol-treated cells. Representative images are shown. The scale bars represent 7 μm. (E) Morphological observation of CRC cells treated with ethanol. (F) Immunofluorescence assay of p-Smad2 protein in ethanol-treated cells or samples from the orthotopic xenograft CRC mouse model. Representative images are shown. The scale bars represent 7 μm. *P < 0.05 vs the control; $P < 0.05 vs 200 EtOH.